Biochemical responses of the aquatic moss Fontinalis antipyretica to Cd,Cu, Pb and Zn determined by chlorophyll fluorescence and protein levels

Biochemical reactions to Cu, Cd, Zn and Pb in the aquatic moss Fontinalis antipyretica were studied in order to characterize the physiological background of the metal response. Chlorophyll fluorescence and intracellular metal localization and stress protein levels were measured. Exposure to 25 or 100 μM Cu over a 7-day period resulted in a decline of chlorophyll fluorescence to about 70% and 52%, respectively. Up to 100 μM Cd caused a decrease in chlorophyll fluorescence to 75%. With all metals used at 25–100 μM concentrations, the intracellular uptake increased. For all metals investigated at 25–100 μM, the intracellular uptake increased. Maximum values were reached at 100 μM Cu, Pb, Zn or Cd exposure. As shown by analytical electron microscopy (EDX, EELS) moss material treated with 50 μM Cu exhibited reduced sulphur levels in the cytoplasm and an increase in phosphate in vacuolar dense particles. EEL-spectra indicated that Cu is chelated in the cytoplasm by SH-groups and coprecipitated with orthophosphate in vacuoles. To monitor the stress response at the protein level, heavy metal induced heat shock protein 70 (hsp70) was measured. An antibody was raised against conserved plant metallothionein p2 motifs derived from Brassica juncea. In all metal-treated samples the antibody bound to proteins of about 8 kDa. However, sequencing failed to reveal significant homologies to known proteins. These experiments provide for the first time results on protein level after heavy metal stress in the aquatic bioindicator moss.